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Eastern equine encephalitis virus in mice I: clinical course and outcome are dependent on route of exposure.

Honnold SP, Mossel EC, Bakken RR, Fisher D, Lind CM, Cohen JW, Eccleston LT, Spurgers KB, Erwin-Cohen R, Bradfute SB, Maheshwari RK, Glass PJ - Virol. J. (2015)

Bottom Line: The majority of those animals exposed by the aerosol route developed severe clinical signs by 4 dpi.Significant differences were also observed in the viral titers of target tissues, with virus being detected in the brain at 6 hpi in the aerosol study.Aerosol exposure to EEEV results in acute onset of clinical signs, rapid neuroinvasion, and 100 % mortality.

View Article: PubMed Central - PubMed

Affiliation: Virology Division, United States Army Medical Research Institute of Infectious Diseases, Fort Detrick, Frederick, MD, 21702, USA. shelley.p.honnold.mil@mail.mil.

ABSTRACT

Background: Eastern equine encephalitis virus (EEEV), an arbovirus, is an important human and veterinary pathogen belonging to one of seven antigenic complexes in the genus Alphavirus, family Togaviridae. EEEV is considered the most deadly of the mosquito-borne alphaviruses due to the high case fatality rate associated with clinical infections, reaching up to 75 % in humans and 90 % in horses. In patients that survive acute infection, neurologic sequelae are often devastating. Although natural infections are acquired by mosquito bite, EEEV is also highly infectious by aerosol. This fact, along with the relative ease of production and stability of this virus, has led it to being identified as a potential agent of bioterrorism.

Methods: To characterize the clinical course and outcome of EEEV strain FL93-939 infection, we compared clinical parameters, cytokine expression, viremia, and viral titers in numerous tissues of mice exposed by various routes. Twelve-week-old female BALB/c mice were infected by the intranasal, aerosol, or subcutaneous route. Mice were monitored for clinical signs of disease and euthanized at specified time points (6 hpi through 8 dpi). Blood and tissues were harvested for cytokine analysis and/or viral titer determination.

Results: Although all groups of animals exhibited similar clinical signs after inoculation, the onset and severity differed. The majority of those animals exposed by the aerosol route developed severe clinical signs by 4 dpi. Significant differences were also observed in the viral titers of target tissues, with virus being detected in the brain at 6 hpi in the aerosol study.

Conclusion: The clinical course and outcome of EEEV infection in mice is dependent on route of exposure. Aerosol exposure to EEEV results in acute onset of clinical signs, rapid neuroinvasion, and 100 % mortality.

No MeSH data available.


Related in: MedlinePlus

Distribution of EEEV FL93-939 in tissues of BALB/c mice exposed by various routes. Shown are the geometric mean virus titer of individual animals in the lung (a), submandibular salivary gland and lymph node (b), spleen (c), and mesenteric lymph node (d) (n = 5/time point). Symbols represent individual animals with values calculated from the geometric mean titer of all dilutions which had at least one visible pfu by standard plaque assay. The mean for the group is shown in the colored dashed line. The limit of detection of the assay is 5 pfu/ml tissue homogenate supernatant.
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Fig6: Distribution of EEEV FL93-939 in tissues of BALB/c mice exposed by various routes. Shown are the geometric mean virus titer of individual animals in the lung (a), submandibular salivary gland and lymph node (b), spleen (c), and mesenteric lymph node (d) (n = 5/time point). Symbols represent individual animals with values calculated from the geometric mean titer of all dilutions which had at least one visible pfu by standard plaque assay. The mean for the group is shown in the colored dashed line. The limit of detection of the assay is 5 pfu/ml tissue homogenate supernatant.

Mentions: Virus titers in the tissues followed similar trends. Although the mice in the IN and AE studies received similar LD50 doses of virus, virus was present in the brain 6 hpi in the AE study, but did not appear until 1 dpi in the IN study (Fig. 5d). Virus titers in the brain continued to increase in both groups, with mice of the AE group consistently having higher titers than mice in the IN group at all time points, peaking with very high titers at 4 dpi in the AE study and 5 dpi in the IN study. In the SC study, virus was first detected in one animal in the brain at 1 dpi, similar to the IN study; however, titers remained lower in the SC study, as compared to the AE and IN studies, for all time points, with virus titer peaking later, at 7 dpi. Virus titers in the lung (Fig. 6a) in the IN and AE studies were similar at 6 hpi, and in both studies continued to rise to fairly high titers through 2 dpi; however, the titers in the AE study were significantly higher than those in the IN study from 2–4 dpi. Virus was not present in the lung in the SC study until 4 dpi and titers remained low even at 6–7 dpi. The mandibular salivary gland and lymph nodes were collected and analyzed together due to their intimate association anatomically. The mandibular lymph nodes, also known as the mandibular and accessory mandibular lymph node, submandibular lymph nodes, or superficial cervical lymph nodes [8], are a small group of lymph nodes that drain the structures of the muzzle [9] and were therefore of interest in the IN and AE studies. As might be expected, virus was present at low levels in the mandibular lymph nodes in both the IN and AE studies at 1 dpi (Fig. 6b) and titers slowly increased throughout the remaining time points, peaking at 5 dpi and 4 dpi, respectively. In the SC study, virus was not present in the mandibular lymph nodes until 4 dpi and remained at relatively low levels through 7 dpi. Virus at this site could be the result of drainage from the nasal cavity or seeding from viremia. Virus titers in the spleen (Fig. 6c) followed similar trends, with virus first appearing in the spleen at 1 dpi in the AE study and at 2 dpi in the IN study; titers increased throughout the study and peaked at 4 dpi and 5 dpi, respectively. In the SC study, virus appeared at 3 dpi, peaked at 4 dpi and slowly decreased over the remaining time points. The mesenteric lymph node, which drains the digestive tract, is a distant lymph node for all routes of infection, and virus present in this lymph node may be indicative of distant viral spread by either blood or lymph. The virus titers in this tissue were low for all routes of infection and peaked at 3 dpi in the AE study, 5 dpi in the IN study, and 7 dpi in the SC study (Fig. 6d). Mean group virus titers of the remaining tissues (liver, heart, kidneys, adrenal glands, and pancreas) evaluated in all three studies are shown in Additional file 1: Figure S1. No virus was present in any of these tissues until 1 dpi and virus titers remained very low throughout most time points. The slight increase in titer in the heart and kidneys could be due to poor perfusion of these vascular organs with PBS prior to tissue collection. Overall, the differences in virus titer in the various tissues are likely due to the variation in delivery method and virus dose. In the IN and AE studies, in which mice received approximately 100LD50, virus was present early on and at high titer in the serum, BAL, brain, and lung. The viral titer in the remaining tissues was lower and peaked later in the time course. While in the SC study, in which mice received approximately 30LD50, the viral titer in tissues was generally lower and peaked later in the course of disease.Fig. 6


Eastern equine encephalitis virus in mice I: clinical course and outcome are dependent on route of exposure.

Honnold SP, Mossel EC, Bakken RR, Fisher D, Lind CM, Cohen JW, Eccleston LT, Spurgers KB, Erwin-Cohen R, Bradfute SB, Maheshwari RK, Glass PJ - Virol. J. (2015)

Distribution of EEEV FL93-939 in tissues of BALB/c mice exposed by various routes. Shown are the geometric mean virus titer of individual animals in the lung (a), submandibular salivary gland and lymph node (b), spleen (c), and mesenteric lymph node (d) (n = 5/time point). Symbols represent individual animals with values calculated from the geometric mean titer of all dilutions which had at least one visible pfu by standard plaque assay. The mean for the group is shown in the colored dashed line. The limit of detection of the assay is 5 pfu/ml tissue homogenate supernatant.
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4588493&req=5

Fig6: Distribution of EEEV FL93-939 in tissues of BALB/c mice exposed by various routes. Shown are the geometric mean virus titer of individual animals in the lung (a), submandibular salivary gland and lymph node (b), spleen (c), and mesenteric lymph node (d) (n = 5/time point). Symbols represent individual animals with values calculated from the geometric mean titer of all dilutions which had at least one visible pfu by standard plaque assay. The mean for the group is shown in the colored dashed line. The limit of detection of the assay is 5 pfu/ml tissue homogenate supernatant.
Mentions: Virus titers in the tissues followed similar trends. Although the mice in the IN and AE studies received similar LD50 doses of virus, virus was present in the brain 6 hpi in the AE study, but did not appear until 1 dpi in the IN study (Fig. 5d). Virus titers in the brain continued to increase in both groups, with mice of the AE group consistently having higher titers than mice in the IN group at all time points, peaking with very high titers at 4 dpi in the AE study and 5 dpi in the IN study. In the SC study, virus was first detected in one animal in the brain at 1 dpi, similar to the IN study; however, titers remained lower in the SC study, as compared to the AE and IN studies, for all time points, with virus titer peaking later, at 7 dpi. Virus titers in the lung (Fig. 6a) in the IN and AE studies were similar at 6 hpi, and in both studies continued to rise to fairly high titers through 2 dpi; however, the titers in the AE study were significantly higher than those in the IN study from 2–4 dpi. Virus was not present in the lung in the SC study until 4 dpi and titers remained low even at 6–7 dpi. The mandibular salivary gland and lymph nodes were collected and analyzed together due to their intimate association anatomically. The mandibular lymph nodes, also known as the mandibular and accessory mandibular lymph node, submandibular lymph nodes, or superficial cervical lymph nodes [8], are a small group of lymph nodes that drain the structures of the muzzle [9] and were therefore of interest in the IN and AE studies. As might be expected, virus was present at low levels in the mandibular lymph nodes in both the IN and AE studies at 1 dpi (Fig. 6b) and titers slowly increased throughout the remaining time points, peaking at 5 dpi and 4 dpi, respectively. In the SC study, virus was not present in the mandibular lymph nodes until 4 dpi and remained at relatively low levels through 7 dpi. Virus at this site could be the result of drainage from the nasal cavity or seeding from viremia. Virus titers in the spleen (Fig. 6c) followed similar trends, with virus first appearing in the spleen at 1 dpi in the AE study and at 2 dpi in the IN study; titers increased throughout the study and peaked at 4 dpi and 5 dpi, respectively. In the SC study, virus appeared at 3 dpi, peaked at 4 dpi and slowly decreased over the remaining time points. The mesenteric lymph node, which drains the digestive tract, is a distant lymph node for all routes of infection, and virus present in this lymph node may be indicative of distant viral spread by either blood or lymph. The virus titers in this tissue were low for all routes of infection and peaked at 3 dpi in the AE study, 5 dpi in the IN study, and 7 dpi in the SC study (Fig. 6d). Mean group virus titers of the remaining tissues (liver, heart, kidneys, adrenal glands, and pancreas) evaluated in all three studies are shown in Additional file 1: Figure S1. No virus was present in any of these tissues until 1 dpi and virus titers remained very low throughout most time points. The slight increase in titer in the heart and kidneys could be due to poor perfusion of these vascular organs with PBS prior to tissue collection. Overall, the differences in virus titer in the various tissues are likely due to the variation in delivery method and virus dose. In the IN and AE studies, in which mice received approximately 100LD50, virus was present early on and at high titer in the serum, BAL, brain, and lung. The viral titer in the remaining tissues was lower and peaked later in the time course. While in the SC study, in which mice received approximately 30LD50, the viral titer in tissues was generally lower and peaked later in the course of disease.Fig. 6

Bottom Line: The majority of those animals exposed by the aerosol route developed severe clinical signs by 4 dpi.Significant differences were also observed in the viral titers of target tissues, with virus being detected in the brain at 6 hpi in the aerosol study.Aerosol exposure to EEEV results in acute onset of clinical signs, rapid neuroinvasion, and 100 % mortality.

View Article: PubMed Central - PubMed

Affiliation: Virology Division, United States Army Medical Research Institute of Infectious Diseases, Fort Detrick, Frederick, MD, 21702, USA. shelley.p.honnold.mil@mail.mil.

ABSTRACT

Background: Eastern equine encephalitis virus (EEEV), an arbovirus, is an important human and veterinary pathogen belonging to one of seven antigenic complexes in the genus Alphavirus, family Togaviridae. EEEV is considered the most deadly of the mosquito-borne alphaviruses due to the high case fatality rate associated with clinical infections, reaching up to 75 % in humans and 90 % in horses. In patients that survive acute infection, neurologic sequelae are often devastating. Although natural infections are acquired by mosquito bite, EEEV is also highly infectious by aerosol. This fact, along with the relative ease of production and stability of this virus, has led it to being identified as a potential agent of bioterrorism.

Methods: To characterize the clinical course and outcome of EEEV strain FL93-939 infection, we compared clinical parameters, cytokine expression, viremia, and viral titers in numerous tissues of mice exposed by various routes. Twelve-week-old female BALB/c mice were infected by the intranasal, aerosol, or subcutaneous route. Mice were monitored for clinical signs of disease and euthanized at specified time points (6 hpi through 8 dpi). Blood and tissues were harvested for cytokine analysis and/or viral titer determination.

Results: Although all groups of animals exhibited similar clinical signs after inoculation, the onset and severity differed. The majority of those animals exposed by the aerosol route developed severe clinical signs by 4 dpi. Significant differences were also observed in the viral titers of target tissues, with virus being detected in the brain at 6 hpi in the aerosol study.

Conclusion: The clinical course and outcome of EEEV infection in mice is dependent on route of exposure. Aerosol exposure to EEEV results in acute onset of clinical signs, rapid neuroinvasion, and 100 % mortality.

No MeSH data available.


Related in: MedlinePlus